Sickle cell disease (SCD) affects approximately 100,000 people in the U.S. but 300,000 babies are born with SCD every year globally. 11% of these children will have a major stroke by age 16, and up to 35% will have a silent stroke, affecting cognitive regions of the brain. Elastic lamina fragmentation in the cerebral arteries were hallmarks identified in autopsy specimens of children with SCD, but mechanisms behind this accelerated arterial remodeling in these children are not known and therefore cannot be prevented. Cysteine cathepsins are powerful proteases implicated in elastin degradation in cardiovascular disease (i.e. atherosclerosis), and our previous studies demonstrated that when peripheral blood mononuclear cells (PBMCs) from people living with SCD adhered to endothelial cells, they significantly increased cathepsin K (catK) activity compared to wildtype PBMCs. This is highly suggestive of a role for cysteine cathepsins in SCD arterial remodeling and strokes, but a major gap remains between proteolytic mechanisms of arterial remodeling and the cellular and inflammatory processes unique to SCD. Our long term goal is to identify therapeutic targets to inhibit proteolytic activity and accelerated elastin degradation in children and adults with SCD. The overall objective of this application is to determine the roles and regulatory mechanisms behind cathepsin mediated arterial remodeling in SCD, and then mitigate arterial damage to prevent strokes. Our central hypothesis is that catK accelerates elastic lamina degradation that precedes strokes in SCD, and catK is induced by inflammatory monocyte signaling, endothelial cells, and interactions between these two via cytokines and microparticle generation. To test this, we will use the Towne's mouse model of sickle cell anemia and the following aims: Aim 1. Determine the role of cathepsins in accelerated elastic lamina degradation due to SCD, and test the efficacy of blocking elastin fragmentation with cathepsin K inhibitors and pathways upstream of cathepsin K expression. Aim 2. Determine cellular mechanisms of SCD-mediated induction of cathepsins in arteries. Aim 3. Determine role of cathepsins in SCD-mediated strokes using assays developed to monitor spontaneous micro-infarcts in young animals using both immunohistochemical methods, but also with the design goal of optimizing 9.4 Tesla MRI scans to quantify stroke lesion number and size. This work is significant because its success will identify novel pharmaceutical treatments to preserve the integrity of carotid and cerebral arteries and to prevent pediatric strokes in people with SCD. Cathepsin inhibitors are already highly sought after products by pharmaceutical companies and identification of cathepsin inhibition as treatment will provide a non-transfusion based drug to prevent SCD strokes in children at risk. Innovative aspects of this project include: 1) Studying arterial complications of SCD, 2) implicating cysteine cathepsins in SCD, a family of enzymes that have been previously unexplored in this context, 3) developing MRI protocols to monitor spontaneous strokes in sickle cell transgenic mice.